From podocyte biology to novel cures for glomerular disease

meeting report

www.kidney-international.org

From podocyte biology to novel cures for glomerular disease Elena Torban1, Fabian Braun2, Nicola Wanner2, Tomoko Takano1, Paul R. Goodyer3, Rachel Lennon4, Pierre Ronco5, Andrey V. Cybulsky1 and Tobias B. Huber2 1

Department of Medicine, McGill University Health Centre Research Institute, McGill University, Montreal, Quebec, Canada; 2III. Department of Medicine, University Medical Center Hamburg-Eppendorf, Hamburg, Germany; 3Department of Pediatrics, McGill University Health Centre Research Institute, McGill University, Montreal, Quebec, Canada; 4Wellcome Centre for Cell-Matrix Research, University of Manchester, Manchester, UK; and 5Sorbonne University, INSERM UMR_S 1155, and Nephrology and Dialysis Department, Hôpital Tenon, Paris France

The podocyte is a key component of the glomerular filtration barrier. Podocyte dysfunction is central to the underlying pathophysiology of many common glomerular diseases, including diabetic nephropathy, glomerulonephritis and genetic forms of nephrotic syndrome. Collectively, these conditions affect millions of people worldwide, and account for the majority of kidney diseases requiring dialysis and transplantation. The 12th International Podocyte Conference was held in Montreal, Canada from May 30 to June 2, 2018. The primary aim of this conference was to bring together nephrologists, clinician scientists, basic scientists and their trainees from all over the world to present their research and to establish networks with the common goal of developing new therapies for glomerular diseases based on the latest advances in podocyte biology. This review briefly highlights recent advances made in understanding podocyte structure and metabolism, experimental systems in which to study podocytes and glomerular disease, disease mediators, genetic and immune origins of glomerulopathies, and the development of novel therapeutic agents to protect podocyte and glomerular injury. Kidney International (2019) 96, 850–861; https://doi.org/10.1016/ j.kint.2019.05.015 Copyright ª 2019, International Society of Nephrology. Published by Elsevier Inc. All rights reserved.

Correspondence: Elena Torban, McGill University, 1001 Decarie Blvd, Montreal, PQ, Canada, H4A 3J1. E-mail: [email protected]; or Tobias B. Huber, University Medical Center Hamburg-Eppendorf, III. Department of Medicine, Martinistr. 52, Hamburg, Germany. E-mail: [email protected] All authors contributed equally. Received 18 February 2019; revised 23 April 2019; accepted 13 May 2019; published online 31 May 2019 850

T

he theme of the 12th International Podocyte Conference (IPC) was “from podocyte biology to novel cures for glomerular disease.” The meeting began with sessions on podocyte development, podocyte ultrastructure, and interactions of podocytes with neighboring cells and the glomerular basement membrane (GBM). Subsequent sessions covered mechanisms of nephrotic syndrome, mediators of podocyte injury, metabolic origins of podocyte disease, genetics, experimental model systems, and therapeutics of glomerulonephritis.

Podocyte origin in development and disease

Our understanding of the molecular and morphological processes underlying glomerular development has improved significantly in recent years. Ultrastructural analysis with cutting-edge electron microscopy approaches has provided insight into the maturation of primitive podocytes and their mature protrusions, as well as the intricate network of foot processes and the slit diaphragm (Figure 1).1 Kann and colleagues2 developed new methods for the analysis of the genetic programs guiding these cellular changes. Using chromatin immunoprecipitation (ChIP) sequencing, they identified a number of genes that are targets of the Wilms Tumor 1 (WT1) transcription factor. In experimental nephrotic syndrome, WT1 binding sites in some genes were lost, while new sites were acquired, implying that WT1-regulated genes may be causative of nephrotic diseases in humans. The tight regulation of the podocyte transcriptome also relies on other mechanisms. Compelling data from Marrone et al.3 were presented on the role of microRNAs (micro RNAs) in podocyte development and disease. This research has revealed specific miRNA clusters involved in the development of nephron progenitor cells. Mutations in the MIR17HG cluster (miR-17-92 in mice) were detected as the first miRNA mutations to cause renal developmental defects in Feingold syndrome, characterized by impaired progenitor cell proliferation and a reduced number of developing nephrons. Conversely, miRNAs might represent a valuable therapeutic option, as overexpression of different miRNA species is associated with modulatory effects on cyst growth in experimental models of polycystic kidney disease and diabetic nephropathy.4 Wanner et al.5 discussed epigenetic control of nephron number in kidney development, a major determinant Kidney International (2019) 96, 850–861

E Torban et al.: Twelfth International Podocyte Conference

Figure 1 | Podocyte origin in development and disease. Nephron development starts with the aggregation of the nephron progenitor cells, initiating the transition from renal vesicle to mature nephron. Podocytes are for the first time detectable in the comma-shaped stage (blue) and later form their characteristic foot processes. In this process, transcription factor WT1, DNA methylation, and microRNA (miRNA) 17 have been shown to play a role. Me, methylation.

of long-term renal function. Nutritional restriction of kidney growth is associated with DNA hypomethylation. DNA methyltransferases Dnmt1 and Dnmt3a are highly expressed in the developing kidney. By analogy to nutritional growth restriction, deletion of Dnmt1 in nephron progenitor cells led to a reduction in nephron number and renal hypoplasia at birth. DNA hypomethylation resulted in downregulation of genes crucial for initiation of nephrogenesis, thus representing a key regulatory event of prenatal renal programming, linking maternal nutritional factors during gestation and reduced nephron number. To further elucidate the relevance of progenitor cells in glomerular development, Lichtnekert et al.6 and Eng et al.7 used elegant fate-tracking experiments to identify cells of renin lineage that exhibit pluripotency and potential to replenish podocytes. An overview was provided of new models of glomerular development, such as those of Takasato et al.8 and Morisane et al.9 in kidney organoids, and podocytes differentiated from induced pluripotent stem cells.10,11 These novel exciting techniques will sharpen our knowledge in the future. The latter offers the potential to model the glomerular filtration barrier on a chip.12 Podocyte architecture: Focus on the cytoskeleton, slit diaphragm, and their regulation in health and disease

Very few cells in higher vertebrates develop a cell body that matches the intricacy of the podocyte, with primary and secondary foot processes forming both a filtration barrier and a signaling hub represented by the slit diaphragm (Figure 2). A significant contributor to this shape and function is the actin cytoskeleton. Rho guanosine triphosphatases (GTPases), with 22 members, have long been known for their involvement in actin cytoskeletal remodeling, cell motility, and podocyte disease. Kidney International (2019) 96, 850–861

meeting report

Gupta et al.13 deepened our understanding of actin dynamics by identifying the Rho guanosine diphosphate dissociation inhibitor, ARHGDIA, as a gene mutated in congenital nephrotic syndrome. At the cellular level, these mutations promote Rac1 hyperactivation,14 which in turn causes podocyte detachment.15 The roles of numerous Rho GTPase regulatory proteins are yet to be elucidated in podocytes, and their roles are actively being investigated. Wegner and colleagues16 identified chloride intracellular channel protein 5a (Clic5a) as an upstream regulator of the actin connecting protein ezrin. Clic5a is expressed in the podocyte apical domain, as is podocalyxin, and controls the activation of phosphatidylinositol-5 kinase and activationspecific phosphorylation of ezrin. The interaction of podocalyxin with the actin cytoskeleton via activated ezrin appears to be vital for intact podocyte structure. Knockout of Clic5a in mice leads to glomerular abnormalities,16 microaneurysms, and hypertension.17 Besides the complex control of the podocyte cytoskeleton, the podocyte connection to the extracellular matrix (ECM), specifically the GBM, plays an essential role in glomerular health. Ishibe et al.18 and Smeeton et al.19 investigated the role of integrins in kidney biology. Deletion of the b1-integrin linking kinase, ILK, in the ureteric bud of mice during kidney development led to reduced branching morphogenesis and severe interstitial fibrosis at 8–10 weeks of age. Similar effects were seen in mice harboring a mutation in the binding site Y783A of talin, a focal adhesion protein connecting the cytoplasmic tail of integrins to the cytoskeleton, and a complete loss of talin led to renal agenesis and neonatal death.20 The crucial slit diaphragm protein nephrin and its phosphorylation have been the focus of research by New and colleagues.21 Disruption of 3 nephrin tyrosine phosphorylation sites (Y3F mice), which serve as adaptor sites for the proteins Nck1 and Nck2, led to proteinuria.21 Likewise, homozygous Y3F/Y3F mice showed a prolonged recovery in the nephrotoxic serum nephritis model, since nephrin clustering and Nck binding were impaired, resulting in decreased nephrin endocytosis and turnover. Nephrin turnover is in part regulated by ShcA, a phosphotyrosine adaptor protein. ShcA associates with multiple phosphorylation sites on nephrin, promotes phosphorylation, and reduces nephrin signaling. Overexpression of ShcA, which is found in several proteinuric kidney diseases, may also reduce nephrin signaling, pointing toward a common pathway involved in the generation or maintenance of proteinuria.22 Mounting evidence suggests that in many glomerulopathies, actin dynamics and cell adhesion are abnormal and lead to subsequent dysregulation of intercellular signaling. Thus, extensive efforts have been made to find therapeutic druggable targets to inhibit and potentially reverse pathogenic changes. A promising candidate is the small molecule Bis-T-23, as presented in work by Schiffer et al.23 and Ono et al.24 Bis-T-23 facilitates the oligomerization of dynamin, a GTPase. Bis-T-23 was found to successfully restore actin polymerization in injured podocytes in several renal disease models by reversing defects in posttranslational protein modification, such as O-GlcNAcylation. 851

meeting report

E Torban et al.: Twelfth International Podocyte Conference

Figure 2 | Podocyte architecture: focus on the cytoskeleton, and the slit diaphragm, and their regulation in health and disease. Rho guanosine triphosphate (GTP)ase pathways, slit diaphragm signaling, and actin interactions, as well as focal adhesions, have been shown to be crucial for podocyte foot process architecture. ARGHDIA, Rho GDP Dissociation Inhibitor Alpha; BIS-T-23, 1,3-Bis(2-Cyano-3-(3,4,5trihydroxy-phenyl)-acrylamino)propane; CLIC5A, Chloride intracellular channel protein 5; NCK1/2, NCK adaptor protein 1/2; P, phosphorylation; RAC1, Rac Family Small GTPase 1; SHCA, Src Homology 2 Domain Containing Adaptor Protein 1.

Podocytes and friends

The podocyte cannot be viewed in isolation when investigating its functional role in health and disease. It has become clear that there is intricate communication between podocytes and other glomerular cells and structures (Figure 3). Randles and colleagues25 defined the composition of the glomerular ECM using proteomic approaches, and these data can now be compared to ECM from other tissues using the powerful Matrisome Project resource (http://matrisomeproject.mit.edu). The glomerular matrisome was compared to cell-derived ECM from podocyte and endothelial cell co-cultures; although there was a significant overlap, key GBM components including collagen IVa3a4a5 have low abundance in vitro.26 To improve understanding of in vivo ECM composition, Lennon and colleagues report a protocol to implant kidney organoids differentiated in vitro from human pluripotent cells into immunodeficient mice to generate perfused glomerular structures with regions of mature GBM and evidence of filtration function.27 Novel imaging techniques have sharpened our understanding of the morphology of ECM proteins. Serial block face-scanning electron microscopy further delineated changes in GBM morphology during the development of Alport syndrome, a hereditary glomerulopathy related to mutations in GBM collagen IV. Changes included subpodocyte expansions of the GBM and podocyte protrusions invading the GBM.28 Beyond that, super-resolution immunofluorescence microscopy opens up entirely new possibilities to investigate the extracellular environment of the podocyte.29 UnnersjöJess et al.30,31 employed similar techniques in combination with hydrogel-based optical clearing to study the morphology and composition of the slit diaphragm in more detail. Concerning cellular crosstalk, Riquier-Brison et al.32 addressed the pathogenic signals that trigger parietal epithelial cell recruitment after podocyte loss, which leads to cell adhesions and sclerosis in the glomerular tuft, i.e., focal segmental glomerulosclerosis (FSGS). In mice infused with 852

angiotensin II, endothelial-specific deletion of the endothelial PAS domain–containing protein 1 (Epas1) gene accentuated albuminuria, recruitment of pathogenic parietal glomerular epithelial cells, and sclerotic lesions. Furthermore, he showed compelling data on the relevance of the tetraspanin CD9 in parietal epithelial cells for crescent formation or the generation of sclerotic lesions in glomerulopathies. Besides this, cells of the macula densa are central in the physiological remodeling of the glomerulus via Wnt signaling and secreted paracrine factors that act on podocyte precursor cells, including cells of the renin lineage. Burger and his group33 have investigated the role of urinary microparticles in glomerular diseases. Specifically, they identified microparticles that are released from cultured podocytes in response to high glucose exposure or mechanical stretch. Microparticles of podocyte origin were present in the

Figure 3 | Podocytes and friends. Podocytes and endothelial cells both contribute to the glomerular basement membrane (GBM). Podocytes, parietal epithelial cells (PECs), and cells from the macula densa communicate via signaling molecules. Urinary microparticles of podocyte origin can lead to downstream signaling in the tubule. Kidney International (2019) 96, 850–861

E Torban et al.: Twelfth International Podocyte Conference

urine of diabetic rats and humans. Furthermore, data from Munkonda et al.34 suggest that podocyte microparticles induce profibrotic signaling in tubular cells. Immune etiology of nephrotic syndrome

Immune cells and immune-epithelial interactions have increasingly become the focus of investigation into the pathogenesis of proteinuric glomerular disease (Figure 4). D’Addio et al.35 introduced the concept of fetomaternal tolerance relying on regulatory T-cells and negative signals through programmed death-ligand 1 (PDL1) costimulation. Using PDL1 blockade, they identified a shift toward IL-17 producing Th17 cells with a decrease in regulatory T-cells leading to a higher abortion rate. This effect could be abrogated by IL-17 neutralization. This research group is involved in the multicenter TANGO study36 aiming at the discovery of prognostic factors for posttransplant recurrence of glomerular diseases with a particular focus on immune factors. Furthermore, they are investigating how changes in the microbiome (by use of dietary modifications) influence subsequent immunologic effects on proteinuria in nephrotic syndrome in children. Colucci presented work from her group on the underlying immunemediated effects in nephrotic syndrome. By examining the effects of rituximab on B- and T-cells in pediatric patients with frequently relapsing or steroid-dependent nephrotic syndrome, they showed that the only denominator of relapse was the reconstitution of memory B-cells independent of the immunosuppressive regime.37 The importance of immune-mediated effects on nephrotic syndrome was further delineated by a genome-wide association study in children with steroidsensitive nephrotic syndrome that identified 3 singlenucleotide polymorphisms in HLA-DQB1, HLA-DRB1, and BTNL2; all of these genetic loci are involved in the immune

Figure 4 | Immune etiology of nephrotic syndrome. Podocytes are affected by circulating factors (such as suPAR), the microbiome or diet, and different immune subpopulations, such as regulatory T cells (Tregs) and Th17 cells. The occurrence of memory B cells can aggravate immune complex depositions. CD8 T cells are able to access podocytes when the Bowman’s capsule is ruptured. PEC, parietal epithelial cell. Kidney International (2019) 96, 850–861

meeting report

response and are associated with independent risk alleles for steroid-sensitive nephrotic syndrome.38 Similarly, Faridi and colleagues39 identified 3 genetic variations in the integrin subunit alpha M gene (encoding CD11b) that are associated with systemic lupus erythematosus. Further research showed that reduction of Toll-like receptor–dependent proinflammatory signals and suppression of interferon-1 signaling was CD11bdependent, indicating that its activation may be a potential therapy in systemic lupus erythematosus. The soluble urokinase-type plasminogen activator receptor (suPAR) has been intensively investigated for its properties as a soluble immune mediator of proteinuric kidney disease.40,41 More recent evidence points toward suPAR being independently associated with chronic kidney disease.42 New studies were presented depicting suPAR as a possible predictor of mortality in type 2 diabetes,43 of estimated glomerular filtration rate decline in cardiovascular disease,44 as well as the progression of chronic kidney disease in children,45 with possible larger implications as a prognostic factor. The direct effects of suPAR on glomerular cells and whether its depletion from patient blood can lead to a robust amelioration of proteinuric kidney diseases are the subject of current studies. An overview was provided of experimental models of podocyte disease, which made note of the fact that models to study alterations in GBM structure in glomerulopathies are lacking. Recent studies in membranous nephropathy46,47 have resulted in the identification of novel nephritogenic antigens and their mediation of podocyte injury, as well as signaling pathways activated by slit diaphragm molecules, including the interaction of nephrin with ephrin-B1.48 The direct cytotoxic effects of immune cells in glomerular injury are supported by an experimental model of rapidly progressive glomerulonephritis. How the microenvironment plays a specific role in the control of immune–epithelial interactions49 was elegantly presented. Enhanced green fluorescent protein (EGFP)-specific CD8þ T-cells from just EGFP death-inducing (Jedi) mice were injected into mice with a podocyte-specific EGFP transgene. In healthy conditions, podocytes were not accessible to cytotoxic T-cells. However, after the disruption of Bowman’s capsule through the induction of nephrotoxic nephritis, the disease phenotype was aggravated by the injection of Jedi cells: mice had higher blood urea nitrogen and urine albumin levels and more severe histologic lesions, including massive depletion of EGFpositive podocytes. EGFP-specific CD8þ T cells were observed near breaches in Bowman’s capsule, suggesting that CD8þ T cells can interact with podocytes only after disruption of Bowman’s capsule, as observed in kidney biopsies from patients with crescentic glomerulonephritis where glomerular infiltration of CD8þ T cells was observed primarily at the site of cellular crescents with rupture of Bowman’s capsule. Detlef Schlöndorff is currently addressing the next questions on the implication of parietal epithelial cells and the role of matrix metalloproteases in this process (Figure 550,51). 853

meeting report

E Torban et al.: Twelfth International Podocyte Conference

Figure 5 | Prof. Detlef Schlöndorff (left) and his son Johannes Schlöndorff at the 12th International Podocyte Conference, Montreal, 2018.

Advances in therapeutics for glomerular nephropathies

In addition to a wide variety of molecular and clinical studies on disease mechanisms, further research has been conducted into novel therapeutic approaches (Figure 6). Results were presented of 2 studies51,52 conducted on potential therapies for FSGS. In recurrent or de novo FSGS resistant to rituximab or therapeutic plasma exchange, adrenocorticotropic hormone gel administration led to a decrease in proteinuria, though to varying degrees.51 A second prospective study concluded that preemptive rituximab administration or plasma exchange did not prevent the recurrence of FSGS after transplantation while remaining a viable therapeutic option after recurrence.52 Work by Harris et al.53 was presented that further delineated the effects of circulating factors in FSGS. Previously, this group demonstrated that vasodilatorstimulated phosphoprotein (VASP) becomes phosphorylated (activated) upon exposure to plasma exchange material from patients with recurrent FSGS.53 This activation was shown to be dependent on protease-activated receptor-1 (PAR1) and leads to podocyte hypermotility in vitro. Current research 854

focuses on the function of a constitutively active PAR1 in vivo, the activity of PAR1 in human FSGS, and how specific human podocin mutations may impair PAR1 trafficking and interactions with the cytoskeleton, thereby leading to FSGS, as well as potential for pharmacologic rescue. Fornoni presented a prospective therapeutic approach to Alport syndrome.54 She demonstrated that mice with deletion of discoidin domain receptor 1 (DDR1) exhibited a slower progression in COL4A3 knockout Alport mice. DDR1 is a receptor tyrosine kinase expressed on the membranes of podocytes, and excessive de novo production of the collagen IVa1a1a2 network in Alport mice can activate DDR1 and cause podocyte lipid accumulation and lipotoxicity. Pharmacologic modulation of DDR1 or lipid accumulation could be a novel therapeutic approach. Rac1 activation results in podocyte damage, and Rac1activating mutations result in sporadic FSGS.55–57 Zhou and colleagues58 sought downstream targets of Rac1 amenable to therapeutic intervention. They identified Rac1-induced activity of the ion channel transient receptor potential canonical (TRPC)5 and subsequent cytoskeletal remodeling to be a Kidney International (2019) 96, 850–861

E Torban et al.: Twelfth International Podocyte Conference

Figure 6 | Advances in therapeutics for glomerular nephropathies. Several approved and potential therapeutic treatments improve proteinuria, podocyte foot process effacement, and glomerular nephropathies. AC1903, N-(2-furanylmethyl)-1(phenylmethyl)-1H-benzimidazol-2-amine; BRAF, B-Raf ProtoOncogene, Serine/Threonine Kinase; DDR1, Discoidin Domain Receptor Tyrosine Kinase 1; GDC-0879, (E)-2,3-Dihydro-5-[1-(2hydroxyethyl)-3-(4-pyridinyl)-1H-pyrazol-4-yl]-1H-inden-1-one oxime; PAR1, Protease-activated receptor-1; RAC1, Rac Family Small GTPase 1; TRPC5, Transient Receptor Potential Cation Channel Subfamily C Member 5; VASP, Vasodilator Stimulated Phosphoprotein.

druggable target of the small molecule AC1903. AC1903 successfully blocked TRPC5 channel activity, attenuating proteinuria in both a rat genetic FSGS model and hypertensive proteinuric kidney disease. Using another approach, which involved screening more than 5000 US Food and Drug Administration–approved drugs, 2 compounds—BRAFV600E inhibitor GDC-0879 and adenylate cyclase agonist forskolin— were identified to promote podocyte survival, revealing the exciting possibility of repurposing established therapeutics for glomerular diseases.59 These compound screenings and targeted approaches can now be complemented by unbiased “omics” analyses using patient material, as was exemplified by the identification of a compartment- and cell type–specific dysregulation of hypoxia-associated gene transcripts through a weighted correlation network analysis of more than 200 renal biopsies with varying chronic kidney disease stages.60 Such studies harbor the potential of adapting compound screens to promising pathways in future studies. Complement-mediated diseases in the glomerulus

In addition to direct antibody or T-cell–epithelial interactions, humoral immune factors, such as the complement system, are important contributors to many glomerulopathies (Figure 7). Successful treatments such as anti-C5 antibody infusion have outlined the potential of complement-based interventions. A review of the work of Langman and Kihr61 was presented on the genetics of complement-regulatory proteins in C3 glomerulopathy (C3G) and atypical hemolytic syndrome (aHUS). Podocytes express several complement receptors, suggesting that aHUS can affect podocytes directly. Kidney International (2019) 96, 850–861

meeting report

Figure 7 | Complement-mediated diseases in the glomerulus. The complement cascade contributes to many glomerulopathies, such as membraneous nephropathy, C3 glomerulopathy, atypical hemolytic uremic syndrome (aHUS), and ischemia reperfusion transplant tolerance. MASP, mannose-associated serine protease; MBL, mannan-binding lectin.

The potential role for complement therapeutics in C3G was highlighted, albeit with a caution that more research is required.61 The investigation of rare genetic variants in aHUS and C3G has in part been hampered by the small size of patient cohorts. Osborne and colleagues62 were able to detect 371 novel rare genetic variants for aHUS and 82 for C3G through the analysis of 13 complement genes in more than 3500 patients. The resulting database has expanded our understanding of the 2 disease entities, with a clear nonrandom distribution of variants over the affected proteins. Beyond these glomerular pathologies with important involvement of the complement cascade, compelling evidence from Ronco and Debiec63 was presented for the involvement of complement in human membranous nephropathy. There is evidence for the activation of the lectin and alternative complement pathways in membranous nephropathy. Analyzing membranous nephropathy patients with unusual progression, Seikrit and colleagues64 identified antibodies to factor H, a regulatory component of the complement system. Nauser and colleagues65 showed the potential of complement inhibition in ischemia–reperfusion injury and organ transplant tolerance. In particular, they showed an abnormal L-fucose pattern that is identified by a C-type lectin, collectin-11, subsequently triggering the complement cascade via the lectin pathway. Inhibition of collectin-11 led to strong protection from ischemia–reperfusion damage, pointing toward new potential mechanisms of reducing ischemia–reperfusion injury after transplantation. Metabolic origin of glomerular disease

The glomerulus represents a demanding microenvironment requiring high metabolic activity in its cellular components (Figure 8). Recent work of Brinkkoetter et al.66, Ising et al.67, Inoki et al.68, and Welsh et al.69 has made significant advances 855

meeting report

E Torban et al.: Twelfth International Podocyte Conference

Figure 8 | Metabolic origin of glomerular disease. Crucial metabolic processes in the podocyte involve mammalian target of rapamycin (mTOR) pathway, autophagy, ubiquitin/proteasome system (UPS), mitochondria, and endoplasmic reticulum (ER). mTORC1, mammalian target of rapamycin complex1; NOX, NADPH oxidase; oxPhos, oxidative phosphorylation; Ub, ubiquitin.

in our understanding of metabolic control in podocyte health and disease. The mammalian target of rapamycin (mTOR) was identified as a primary regulator of podocyte autophagy during aging and an important signal in guiding compensatory hypertrophy. mTOR dysfunction may contribute to podocyte loss. Their focus has now shifted to mechanisms of energy consumption and oxidative phosphorylation in the podocyte, with new data pointing toward podocyte-specific mechanisms in glucose metabolism. An excess of reactive oxygen species production through reduced nicotinamide adenine dinucleotide phosphate oxidases (NOX) poses a severe, but potentially druggable, threat to the podocyte, as shown by Chris Kennedy’s group70: Both expression of NOX5 in mice and overexpression of NOX4 in a diabetic mouse model resulted in albuminuria and podocyte damage, and inhibition of the latter through knockout or pharmacologic intervention ameliorated the disease phenotype in diabetes.70–73 Compelling data were presented implicating calcium channels TRPC5 and TRPC6 in this process.72 Besides energy metabolism, Andrey Bartram and colleagues74 delineated the pathologic impact of endoplasmic reticulum (ER) stress and proteotoxicity on podocytes. It has been appreciated that mutations in a-actinin-4 result in a genetic form of FSGS. One reason for the occurrence of podocyte damage is proteotoxicity and aggregate formation. These effects can be ameliorated by administering the chemical chaperone 4-phenyl butyric acid to mice with FSGS associated with expression of mutant a-actinin-4. The chemical chaperone reduces enhanced protein misfolding and ER stress.75 The role for ER stress was also demonstrated by a podocyte-specific knockout of inositol-requiring enzyme-1a (IRE1a; an ER transmembrane protein and transducer of ER stress), which resulted in albuminuria and foot-process effacement in aging mice, and was at least in part related to impaired autophagy.76 Knockout of IRE1a also exacerbated injury in anti-GBM nephritis. Endocytosis and recycling of proteins in podocytes (i.e., nephrin) is deregulated in disease states, such as diabetes.77 Beeken and colleagues78 have further investigated the role of podocyte proteostasis by examining the 2 main protein degradation mechanisms—the 856

ubiquitin-proteasome system (UPS) and autophagy. Both systems vary in their activity in different glomerular diseases. In fact, increases in specific UPS proteins can differentiate between minimal change disease and FSGS. Impaired autophagy through knockout of the autophagy protein 5 (ATG5) gene in podocytes leads to proteinuria and renal insufficiency.79 Tampering with the UPS likewise results in proteinuria and exacerbation of injury in experimental models of nephritis, underlining the importance of tightly regulated protein metabolism in sustaining podocyte function.80,81 Genetics of glomerular disease

The study of genetics in glomerular diseases has had a major impact on the understanding of podocyte biology (Figure 9). The field is rapidly expanding with the identification of more and more pathogenic genetic variants resulting in a disease phenotype. COL4A3/4/5 mutations were previously thought to be associated exclusively with Alport syndrome. Barua et al.82 elaborated on the importance of exome sequencing in patient families exhibiting proteinuric kidney disease and FSGS by showing that specific COL4A3/4/5 mutations are present in a significant proportion of these families. The potential wider

Figure 9 | Genetics of glomerular disease. Novel mutations causing monogenetic nephrotic diseases have been found. The effect of microRNA (miRNA) on podocyte biology is under investigation. Genome-wide association studies (GWAS) detect correlations between single-nucleotide polymorphisms (SNPs) and disease. Single-cell RNA-sequencing (scRNA-seq) elucidates gene expression in single glomerular cells. Kidney International (2019) 96, 850–861

meeting report

E Torban et al.: Twelfth International Podocyte Conference

importance of Alport gene mutations was highlighted in a report of recent findings from a large genome-wide association study in diabetic kidney disease. One significant locus was a variant in COL4A3.83 Spearheading the search for new genes in steroid-resistant nephrotic syndrome (SRNS), Sadowski et al.84 have continued to broaden our understanding of genetic causes of kidney disease, demonstrating that close to a third of familial SRNS cases are monogenic diseases. New mutations have been discovered in DNA-damage response complexes,85 sphingosine metabolism,86 regulation of small GTPases,87 and nucleoporins.88 A technique that has been changing the landscape of genetic research over the last year has been single-cell RNA sequencing (scRNAseq). The first atlas of scRNAseq of the murine kidney was recently published.89 The dataset represents a valuable resource for the correlation of past, current, and future genome-wide association study datasets, as singlenucleotide polymorphisms can be ascribed to the cells with the highest expression of the host gene. A presentation was made on how a monogenic disease can increase the understanding of podocyte biology using the example of Fabry disease. Previous studies underlined the impact of dysregulated autophagy, profibrotic signaling, and deranged lipid metabolism on podocyte health.90–92 Jarad and colleagues93 investigated metabolic and genetic modifiers in Alport syndrome and discovered that albumin and its filtration through the damaged GBM is a significant contributor to the disease phenotype. Furthermore, others showed that silencing of the microRNA-21 by specific oligonucleotides led to decreased fibrogenesis as a result of enhanced peroxisome proliferator-activated receptor-a/retinoic X receptor activity and improved mitochondrial function, possibly opening an opportunity for therapy.94 Similarly, inducible expression of a COL4A3 transgene in mice, which leads to secretion and assembly of collagen IV a3a4a5

heterotrimers by podocytes, ameliorated the Alport phenotype by restoring GBM integrity.95 This group also uncovered genetic modifiers of the disease course, such as a human mutation in LAMB2.96 Model systems to study podocytes

Podocyte research relies heavily on model systems. A dedicated session, therefore, highlighted the advances in establishing novel models for glomerular disease and their analysis (Figure 10). Siegerist and colleagues97 introduced superresolution microscopy and the measurement of slit diaphragm length per glomerular capillary surface as a novel tool to robustly evaluate foot process effacement. The architecture of the Drosophila nephrocyte resembles the structure of the podocyte foot process and slit diaphragm in striking ways. Accordingly, this Drosophila model has emerged as a valuable research tool for the investigation of podocyte biology. One presentation focused on 2 studies in nephrocytes98,99 that revealed, respectively, the importance of small GTPases Rab 5, 7, and 11, and of coenzyme Q10 in nephrocyte health, pointing to similar functions of these molecules in mammalian podocyte biology. Vineet Gupta’s group established a novel highthroughput assay to simultaneously screen for the effects of multiple pharmacologic compounds on blocking drug-induced podocyte toxicity (seen as changes in the integrity of actin cytoskeleton and focal adhesions) using podocytes grown in a multiwell dish. Lee and coworkers were able to identify 1% of more than 2000 US Food and Drug Administration–approved compounds to be protective in podocytes; one of these compounds, pyrintegrin, showed similar protective effects in vivo.100 Since then, the procedure has been fully automated, enabling enhanced screening of more compounds. A follow-up study indicates that the slit guidance ligand 2 (SLIT2)/roundabout guidance receptor 2 (ROBO2)/SLIT-ROBO Rho GTPase

Figure 10 | Model systems to study podocytes. Novel methods and models are being employed to uncover podocyte function. Superresolution microscopy has greatly improved resolution of slit diaphragm stainings. High-throughput screening assays of US Food and Drug Administration–approved drug libraries uncover podocyte protective compounds. Drosophila melanogasternephrocytes function as simplified podocyte models. Humanized mouse models are a valuable tool to delineate immune processes mediated by mononuclear cells in patients. Kidney International (2019) 96, 850–861

857

meeting report

Figure 11 | Scanning electron microscopic picture of podocytes with pseudocoloring of foot processes (original picture from Martin Helmstädter and Tobias B. Huber).

activating protein 1/non-muscle myosin IIA heavy chain axis plays a role in podocyte adhesion. Interference with ROBO2 signaling has been proposed as a potential therapeutic option in glomerular diseases.101 The investigation of immune–epithelial interactions poses specific problems when studying the process in rodent models, as the current inbred strains only partially resemble the human immune system. Hahm and her group41 examined the potential of humanized mice by injecting peripheral blood mononuclear cells into immunodeficient mice. Indeed, they showed that, for example, peripheral blood mononuclear cells from patients with recurrent FSGS could engraft in the new host body and trigger immune responses such as suPAR release and foot process effacement. These responses were abrogated upon transplantation of a peripheral blood mononuclear cell population depleted of CD34 cells.41 Additional data were presented on how the humanized mouse model represents a valuable tool to delineate immune processes mediated by mononuclear cells in patients. Controversies and discussions

Some of the presentations at the IPC and the associated discussions raised controversies. For example, strong indirect evidence supports an extrarenal cause for idiopathic FSGS. It has been proposed that a circulating factor toxic to podocytes is produced by the cells of a dysregulated immune system; however, after a number of studies, the factor’s identity and the specific cell lineage producing it remain elusive. This may not be surprising; although we typically refer to FSGS as a single type of podocyte glomerulopathy, FSGS represents a histologic pattern, most likely with distinct etiologies. Indeed, at the IPC, it was highlighted that mutations in the COL4A3/ 4/5 genes may be a substantial and underappreciated cause of 858

E Torban et al.: Twelfth International Podocyte Conference

Figure 12 | Podocyte diseases, cell–cell interactions, cellular targets, methods, and model. CKD, chronic kidney disease; DKD, diabetic kidney disease; DNAm, DNA methylation; ECM, extracellular matrix; FSGS, focal segmental glomerular sclerosis; GBM, glomerular basement membrane; iPSCs, induced pluripotent stem cells; MN, membranous nephropathy; PECs, parietal epithelial cells; RPGN, rapid progressive glomerulonephropathy; scRNA-seq, single cell RNA sequencing; TFs, transcription factors.

FSGS. Likewise, so-called “idiopathic” FSGS may have heterogenous pathophysiology. Serum levels of suPAR appear to be strong predictors of declining renal function and cardiovascular disease,42 but whether suPAR is the driver of chronic kidney disease remains controversial, and especially, if it functions as the circulating podocyte-toxic factor.40,102,103 Clinical trials testing the effects of suPAR absorption therapy may be able to resolve this debate. The lack of clear appreciation of the heterogeneity in FSGS may also be responsible for conflicting results of FSGS treatments presented at the IPC. It remains unclear whether drugs used to treat steroidresponsive nephrotic syndrome (rituximab, glucocorticoids, calcineurin inhibitors, and others) have any role in treating FSGS that recurs after kidney transplantation. Putative downstream mediators of podocyte-toxic factor(s) were presented at the IPC, including PAR1, b3-integrin, dynamin, and other molecules. Participants of the IPC debated whether TRPC5 or TRPC6 may drive podocyte injury.58,104,105 Studies of TRPC isoform-specific inhibitors in patient-derived podocytes may be a way to resolve this controversy. Discussions at the IPC identified several obstacles to progress in understanding podocyte biology and disease. Genetic studies have focused on defining pathogenic monogenic mutations responsible for specific podocytopathies. Mutations in complement regulatory proteins are believed to contribute to the pathogenesis of aHUS and C3 glomerulopathy. However, it was noted at the IPC that such mutations may be more widespread (e.g., in membranous nephropathy), which raises the need for examining mutations across several diseases to draw proper conclusions on genotypes and phenotypes. The Kidney International (2019) 96, 850–861

E Torban et al.: Twelfth International Podocyte Conference

majority of the research efforts have been centered on podocytes themselves, and there is a paucity of information on other cells that have an impact on podocyte function. For example, the contribution of endothelium or parietal cells to specific forms of glomerulopathies is unclear, and detailed phenotypes of patients’ immune cells are not known. Such knowledge is essential to improve and refine therapeutic approaches, e.g., anti-complement therapy in aHUS. It is recognized that metabolism in cultured cells tends to be glycolytic while mitochondria provide energy in vivo. Surprisingly, data presented at the IPC supported a major role for glycolysis in podocytes in vivo, although it remains to be determined if mitochondrial adenosine triphosphate production is dispensable in health and in glomerulopathies. Potential effects of the environment on the function of immune cells were noted at the IPC, but at present, such effects are considered infrequently in experimental studies on glomerular disease. Likewise, standardized and reproducible experimental models to study podocyte injury are lacking. In this regard, humanized rodent models and “mini” organoids from patient-derived cells will likely provide a new direction, although further fine-tuning of these emerging models is required to determine the extent to which they recapitulate the functions of podocytes or immune cells in human health and disease (Figure 11). Conclusion

The 2018 12th International Podocyte Conference in Montreal highlighted the most recent discoveries in podocyte biology and mechanisms of proteinuric disease. It brought together various stakeholders, including clinicians, scientists, and trainees from academia and the pharmaceutical industry, and created a sense of excitement regarding an increasingly prolific pace of discovery in the field. The conference brought to light developments in transcriptional and epigenetic control of podocyte gene expression. There were new insights into the dynamics of the slit diaphragm, actin cytoskeleton, and regulators, including Rho GTPases. The importance of crosstalk of podocytes with other glomerular cells and GBM was emphasized. The roles of immune mediators, complement, protein folding, and regulation of proteostasis in podocyte diseases were highlighted. Finally, novel techniques in imaging and scRNAseq were presented. Several regulators and pathways may constitute druggable targets for podocyte diseases, and various promising therapeutic approaches were discussed (Figure 12). These presentations provide hope that, in the future, podocyte diseases will be preventable or attenuated in many patients by use of such mechanism-based therapies. DISCLOSURE

All the authors declared no competing interests. REFERENCES 1. Ichimura K, Kakuta S, Kawasaki Y, et al. Morphological process of podocyte development revealed by block-face scanning electron microscopy. J Cell Sci. 2017;130:132–142. Kidney International (2019) 96, 850–861

meeting report

2. Kann M, Ettou S, Jung YL, et al. Genome-wide analysis of Wilms’ tumor 1-controlled gene expression in podocytes reveals key regulatory mechanisms. J Am Soc Nephrol. 2015;26:2097–2104. 3. Marrone AK, Stolz DB, Bastacky SI, et al. MicroRNA-17w92 is required for nephrogenesis and renal function. J Am Soc Nephrol. 2014;25:1440–1452. 4. Pandey P, Qin S, Ho J, et al. Systems biology approach to identify transcriptome reprogramming and candidate microRNA targets during the progression of polycystic kidney disease. BMC Syst Biol. 2011;5:56. 5. Wanner N, Vornweg J, Combes A, et al. DNA methyltransferase 1 controls nephron progenitor cell renewal and differentiation. J Am Soc Nephrol. 2019;30:63–78. 6. Lichtnekert J, Kaverina NV, Eng DG, et al. Renin-angiotensinaldosterone system inhibition increases podocyte derivation from cells of renin lineage. J Am Soc Nephrol. 2016;27:3611–3627. 7. Eng DG, Kaverina NV, Schneider RRS, et al. Detection of renin lineage cell transdifferentiation to podocytes in the kidney glomerulus with dual lineage tracing. Kidney Int. 2018;93:1240–1246. 8. Takasato M, Er PX, Chiu HS, et al. Kidney organoids from human iPS cells contain multiple lineages and model human nephrogenesis. Nature. 2015;526:564–568. 9. Morizane R, Lam AQ, Freedman BS, et al. Nephron organoids derived from human pluripotent stem cells model kidney development and injury. Nature Biotechnol. 2015;33:1193–1200. 10. Yoshimura Y, Taguchi A, Tanigawa S, et al. Manipulation of nephronpatterning signals enables selective induction of podocytes from human pluripotent stem cells. J Am Soc Nephrol. 2019. ASN. 2018070747–18. 11. Nishinakamura R. The era of human developmental nephrology. J Am Soc Nephrol. 2018;29:705–706. 12. Musah S, Mammoto A, Ferrante TC, et al. Mature induced-pluripotentstem-cell-derived human podocytes reconstitute kidney glomerularcapillary-wall function on a chip. Nat Biomed Eng. 2017;1:0069–12. 13. Gupta IR, Baldwin C, Auguste D, et al. ARHGDIA: a novel gene implicated in nephrotic syndrome. J Med Genet. 2013;50:330–338. 14. Auguste D, Maier M, Baldwin C, et al. Disease-causing mutations of RhoGDIa induce Rac1 hyperactivation in podocytes. Small GTPases. 2016;7:107–121. 15. Robins R, Baldwin C, Aoudjit L, et al. Rac1 activation in podocytes induces the spectrum of nephrotic syndrome. Kidney Int. 2017;92:349–364. 16. Wegner B, Al-Momany A, Kulak SC, et al. CLIC5A, a component of the ezrin-podocalyxin complex in glomeruli, is a determinant of podocyte integrity. Am J Physiol Renal Physiol. 2010;298:F1492–F1503. 17. Tavasoli M, Al-Momany A, Wang X, et al. Both CLIC4 and CLIC5A activate ERM proteins in glomerular endothelium. Am J Physiol Renal Physiol. 2016;311:F945–F957. 18. Ishibe S, Karihaloo A, Ma H, et al. Met and the epidermal growth factor receptor act cooperatively to regulate final nephron number and maintain collecting duct morphology. Development. 2009;136: 337–345. 19. Smeeton J, Zhang X, Bulus N, et al. Integrin-linked kinase regulates p38 MAPK-dependent cell cycle arrest in ureteric bud development. Development. 2010;137:3233–3243. 20. Mathew S, Palamuttam RJ, Mernaugh G, et al. Talin regulates integrin b1-dependent and -independent cell functions in ureteric bud development. Development. 2017;144:4148–4158. 21. New LA, Martin CE, Scott RP, et al. Nephrin tyrosine phosphorylation is required to stabilize and restore podocyte foot process architecture. J Am Soc Nephrol. 2016;27:2422–2435. 22. Martin CE, Petersen KA, Aoudjit L, et al. ShcA adaptor protein promotes nephrin endocytosis and is upregulated in proteinuric nephropathies. J Am Soc Nephrol. 2018;29:92–103. 23. Schiffer M, Teng B, Gu C, et al. Pharmacological targeting of actindependent dynamin oligomerization ameliorates chronic kidney disease in diverse animal models. Nat Med. 2015;21:601–609. 24. Ono S, Kume S, Yasuda-Yamahara M, et al. O-linked b-Nacetylglucosamine modification of proteins is essential for foot process maturation and survival in podocytes. Nephrol Dial Transplant. 2017;32: 1477–1487. 25. Randles MJ, Humphries MJ, Lennon R. Proteomic definitions of basement membrane composition in health and disease. Matrix Biol. 2017;57-58:12–28. 26. Byron A, Randles MJ, Humphries JD, et al. Glomerular cell cross-talk influences composition and assembly of extracellular matrix. J Am Soc Nephrol. 2014;25:953–966.

859

meeting report

27. Bantounas I, Ranjzad P, Tengku F, et al. Generation of functioning nephrons by implanting human pluripotent stem cell-derived kidney progenitors. Stem Cell Rep. 2018;10:766–779. 28. Randles MJ, Collinson S, Starborg T, et al. Three-dimensional electron microscopy reveals the evolution of glomerular barrier injury. Sci Rep. 2016;6:35068. 29. Suleiman H, Zhang L, Roth R, et al. Nanoscale protein architecture of the kidney glomerular basement membrane. Elife. 2013;2:e01149. 30. Unnersjö-Jess D, Scott L, Blom H, et al. Super-resolution stimulated emission depletion imaging of slit diaphragm proteins in optically cleared kidney tissue. Kidney Int. 2015:1–5. 31. Unnersjö-Jess D, Scott L, Sevilla SZ, et al. Confocal super-resolution imaging of the glomerular filtration barrier enabled by tissue expansion. Kidney Int. 2018;93:1008–1013. 32. Riquier-Brison ADM, Sipos A, Prokai A, et al. The macula densa prorenin receptor is essential in renin release and blood pressure control. Am J Physiol Renal Physiol. 2018;315:F521–F534. 33. Burger D, Thibodeau J-F, Holterman CE, et al. Urinary podocyte microparticles identify prealbuminuric diabetic glomerular injury. J Am Soc Nephrol. 2014;25:1401–1407. 34. Munkonda MN, Akbari S, Landry C, et al. Podocyte-derived microparticles promote proximal tubule fibrotic signaling via p38 MAPK and CD36. J Extracell Vesicles. 2018;7:1432206. 35. D’Addio F, Riella LV, Mfarrej BG, et al. The link between the PDL1 costimulatory pathway and Th17 in fetomaternal tolerance. J Immunol. 2011;187:4530–4541. 36. Uffing A, Pérez-Sáez MJ, La Manna G, et al. A large, international study on post-transplant glomerular diseases: the TANGO project. BMC Nephrol. 2018;19:229. 37. Colucci M, Carsetti R, Cascioli S, et al. B cell reconstitution after rituximab treatment in idiopathic nephrotic syndrome. J Am Soc Nephrol. 2016;27:1811–1822. 38. Debiec H, Dossier C, Letouzé E, et al. Transethnic, genome-wide analysis reveals immune-related risk alleles and phenotypic correlates in pediatric steroid-sensitive nephrotic syndrome. J Am Soc Nephrol. 2018;29:2000–2013. 39. Faridi MH, Khan SQ, Zhao W, et al. CD11b activation suppresses TLRdependent inflammation and autoimmunity in systemic lupus erythematosus. J Clin Invest. 2017;127:1271–1283. 40. Wei C, Hindi El S, Li J, et al. Circulating urokinase receptor as a cause of focal segmental glomerulosclerosis. Nat Med. 2011;17:952–960. 41. Hahm E, Wei C, Fernandez I, et al. Bone marrow-derived immature myeloid cells are a main source of circulating suPAR contributing to proteinuric kidney disease. Nat Med. 2017;23:100–106. 42. Hayek SS, Sever S, Ko Y-A, et al. Soluble urokinase receptor and chronic kidney disease. N Engl J Med. 2015;373:1916–1925. 43. Hayek SS, Divers J, Raad M, et al. Predicting mortality in African Americans with type 2 diabetes mellitus: soluble urokinase plasminogen activator receptor, coronary artery calcium, and high-sensitivity C-reactive protein. J Am Heart Assoc. 2018;7: pii: e008194. 44. Hayek SS, Ko Y-A, Awad M, et al. Cardiovascular disease biomarkers and suPAR in predicting decline in renal function: a prospective cohort study. Kidney Int Rep. 2017;2:425–432. 45. Schaefer F, Trachtman H, Wühl E, et al. Association of serum soluble urokinase receptor levels with progression of kidney disease in children. JAMA Pediatr. 2017;171:e172914–e172914. 46. Tomas NM, Hoxha E, Reinicke AT, et al. Autoantibodies against thrombospondin type 1 domain-containing 7A induce membranous nephropathy. J Clin Invest. 2016;126:2519–2532. 47. Tomas NM, Meyer-Schwesinger C, Spiegel von H, et al. A heterologous model of thrombospondin type 1 domain-containing 7A-associated membranous nephropathy. J Am Soc Nephrol. 2017;28:3262–3277. 48. Fukusumi Y, Zhang Y, Yamagishi R, et al. Nephrin-binding ephrin-B1 at the slit diaphragm controls podocyte function through the JNK pathway. J Am Soc Nephrol. 2018:ASN.2017090993. 49. Chen A, Lee K, D’Agati VD, et al. Bowman’s capsule provides a protective niche for podocytes from cytotoxic CD8þ T cells. J Clin Invest. 2018;128:3413–3424. 50. Schlondorff D. Putting the glomerulus back together: per aspera ad astra (“a rough road leads to the stars”). Kidney Int. 2014;85:991–998. 51. Alhamad T, Manllo Dieck J, Younus U, et al. ACTH gel in resistant focal segmental glomerulosclerosis after kidney transplantation. Transplantation. 2019;103:202–209.

860

E Torban et al.: Twelfth International Podocyte Conference

52. Alasfar S, Matar D, Montgomery RA, et al. Rituximab and therapeutic plasma exchange in recurrent focal segmental glomerulosclerosis postkidney transplantation. Transplantation. 2018;102:e115–e120. 53. Harris JJ, McCarthy HJ, Ni L, et al. Active proteases in nephrotic plasma lead to a podocin-dependent phosphorylation of VASP in podocytes via protease activated receptor-1. J Pathol. 2013;229: 660–671. 54. Richter H, Satz AL, Bedoucha M, et al. DNA-encoded library-derived DDR1 inhibitor prevents fibrosis and renal function loss in a genetic mouse model of Alport Syndrome. ACS Chem Biol. 2019;14: 37–49. 55. Akilesh S, Suleiman H, Yu H, et al. Arhgap24 inactivates Rac1 in mouse podocytes, and a mutant form is associated with familial focal segmental glomerulosclerosis. J Clin Invest. 2011;121:4127– 4137. 56. Gee HY, Saisawat P, Ashraf S, et al. ARHGDIA mutations cause nephrotic syndrome via defective RHO GTPase signaling. J Clin Invest. 2013;123: 3243–3253. 57. Yu H, Artomov M, Brähler S, et al. A role for genetic susceptibility in sporadic focal segmental glomerulosclerosis. J Clin Invest. 2016;126: 1603. 58. Zhou Y, Castonguay P, Sidhom E-H, et al. A small-molecule inhibitor of TRPC5 ion channels suppresses progressive kidney disease in animal models. Science. 2017;358:1332–1336. 59. Sieber J, Wieder N, Clark A, et al. GDC-0879, a BRAFV600E inhibitor, protects kidney podocytes from death. Cell Chem Biol. 2018;25:175–184. e4. 60. Shved N, Warsow G, Eichinger F, et al. Transcriptome-based network analysis reveals renal cell type-specific dysregulation of hypoxiaassociated transcripts. Sci Rep. 2017;7:8576. 61. Keir LS, Langman CB. Complement and the kidney in the setting of Shiga-toxin hemolytic uremic syndrome, organ transplantation, and C3 glomerulonephritis. Transfus Apher Sci. 2016;54:203–211. 62. Osborne AJ, Breno M, Borsa NG, et al. Statistical validation of rare complement variants provides insights into the molecular basis of atypical hemolytic uremic syndrome and C3 glomerulopathy. J Immunol. 2018;200:2464–2478. 63. Ronco P, Debiec H. A podocyte view of membranous nephropathy: from Heymann nephritis to the childhood human disease. Pflugers Arch. 2017;469:997–1005. 64. Seikrit C, Ronco P, Debiec H. Factor H autoantibodies and membranous nephropathy. N Engl J Med. 2018;379:2479–2481. 65. Nauser CL, Howard MC, Fanelli G, et al. Collectin-11 (CL-11) is a major sentinel at epithelial surfaces and key pattern recognition molecule in complement-mediated ischaemic injury. Front Immunol. 2018;9: 2023. 66. Brinkkoetter PT, Bork T, Salou S, et al. Anaerobic glycolysis maintains the glomerular filtration barrier independent of mitochondrial metabolism and dynamics. Cell Rep. 2019;27:1551–1566.e5. 67. Ising C, Koehler S, Brähler S, et al. Inhibition of insulin/IGF-1 receptor signaling protects from mitochondria-mediated kidney failure. EMBO Mol Med. 2015;7:275–287. 68. Inoki K, Mori H, Wang J, et al. mTORC1 activation in podocytes is a critical step in the development of diabetic nephropathy in mice. J Clin Invest. 2011;121:2181–2196. 69. Welsh GI, Hale LJ, Eremina V, et al. Insulin signaling to the glomerular podocyte is critical for normal kidney function. Cell Metab. 2010;12:329– 340. 70. Sedeek M, Gutsol A, Montezano AC, et al. Renoprotective effects of a novel Nox1/4 inhibitor in a mouse model of Type 2 diabetes. Clin Sci. 2013;124:191–202. 71. Holterman CE, Thibodeau J-F, Towaij C, et al. Nephropathy and elevated BP in mice with podocyte-specific NADPH oxidase 5 expression. J Am Soc Nephrol. 2014;25:784–797. 72. Ilatovskaya DV, Blass G, Palygin O, et al. A NOX4/TRPC6 pathway in podocyte calcium regulation and renal damage in diabetic kidney disease. J Am Soc Nephrol. 2018;29:1917–1927. 73. Cowley AW, Yang C, Zheleznova NN, et al. Evidence of the importance of Nox4 in production of hypertension in Dahl salt-sensitive rats. Hypertension. 2016;67:440–450. 74. Bartram MP, Habbig S, Pahmeyer C, et al. Three-layered proteomic characterization of a novel ACTN4 mutation unravels its pathogenic potential in FSGS. Hum Mol Genet. 2016;25:1152–1164. Kidney International (2019) 96, 850–861

E Torban et al.: Twelfth International Podocyte Conference

75. Yee A, Papillon J, Guillemette J, et al. Proteostasis as a therapeutic target in glomerular injury associated with mutant a-actinin-4. Am J Physiol Renal Physiol. 2018;315:F954–F966. 76. Kaufman DR, Papillon J, Larose L, et al. Deletion of inositol-requiring enzyme-1a in podocytes disrupts glomerular capillary integrity and autophagy. Mol Biol Cell. 2017;28:1636–1651. 77. Dumont V, Tolvanen TA, Kuusela S, et al. PACSIN2 accelerates nephrin trafficking and is up-regulated in diabetic kidney disease. FASEB J. 2017;31:3978–3990. 78. Beeken M, Lindenmeyer MT, Blattner SM, et al. Alterations in the ubiquitin proteasome system in persistent but not reversible proteinuric diseases. J Am Soc Nephrol. 2014;25:2511–2525. 79. Hartleben B, Gödel M, Meyer-Schwesinger C, et al. Autophagy influences glomerular disease susceptibility and maintains podocyte homeostasis in aging mice. J Clin Invest. 2010;120:1084–1096. 80. Radón V, Czesla M, Reichelt J, et al. Ubiquitin C-terminal hydrolase L1 is required for regulated protein degradation through the ubiquitin proteasome system in kidney. Kidney Int. 2018;93:110–127. 81. Lohmann F, Sachs M, Meyer TN, et al. UCH-L1 induces podocyte hypertrophy in membranous nephropathy by protein accumulation. Biochim Biophys Acta. 2014;1842:945–958. 82. Barua M, John R, Stella L, et al. X-linked glomerulopathy due to COL4A5 founder variant. Am J Kidney Dis. 2018;71:441–445. 83. Salem R, Todd JN, Sandholm N, et al. Genome-wide association study of diabetic kidney disease highlights biology involved in renal basement membrane collagen. bioRxiv. 2018:499616. 84. Sadowski CE, Lovric S, Ashraf S, et al. A single-gene cause in 29.5% of cases of steroid-resistant nephrotic syndrome. J Am Soc Nephrol. 2015;26:1279–1289. 85. Braun DA, Rao J, Mollet G, et al. Mutations in KEOPS-complex genes cause nephrotic syndrome with primary microcephaly. Nat Genet. 2017;49:1529–1538. 86. Lovric S, Goncalves S, Gee HY, et al. Mutations in sphingosine-1phosphate lyase cause nephrosis with ichthyosis and adrenal insufficiency. J Clin Invest. 2017;127:912–928. 87. Ashraf S, Kudo H, Rao J, et al. Mutations in six nephrosis genes delineate a pathogenic pathway amenable to treatment. Nat Commun. 2018;9: 1960. 88. Braun DA, Sadowski CE, Kohl S, et al. Mutations in nuclear pore genes NUP93, NUP205 and XPO5 cause steroid-resistant nephrotic syndrome. Nat. Genet. 2016;48:457–465. 89. Park J, Shrestha R, Qiu C, et al. Single-cell transcriptomics of the mouse kidney reveals potential cellular targets of kidney disease. Science. 2018: eaar2131.

Kidney International (2019) 96, 850–861

meeting report

90. Liebau MC, Braun F, Höpker K, et al. Dysregulated autophagy contributes to podocyte damage in Fabry’s disease. PLoS ONE. 2013;8: e63506. 91. Sanchez-Niño MD, Carpio D, Sanz AB, et al. Lyso-Gb3 activates Notch1 in human podocytes. Hum Mol Genet. 2015;24:5720–5732. 92. Najafian B, Svarstad E, Bostad L, et al. Progressive podocyte injury and globotriaosylceramide (GL-3) accumulation in young patients with Fabry disease. Kidney Int. 2010;79:663–670. 93. Jarad G, Knutsen RH, Mecham RP, et al. Albumin contributes to kidney disease progression in Alport syndrome. AM J Physiol Renal Physiol. 2016;311:F120–F130. 94. Gomez IG, MacKenna DA, Johnson BG, et al. Anti-microRNA-21 oligonucleotides prevent Alport nephropathy progression by stimulating metabolic pathways. J Clin Invest. 2015;125:141–156. 95. Lin X, Suh JH, Go G, et al. Feasibility of repairing glomerular basement membrane defects in Alport syndrome. J Am Soc Nephrol. 2014;25:687–692. 96. Funk SD, Bayer RH, Malone AF, et al. Pathogenicity of a human laminin b2 mutation revealed in models of Alport Syndrome. J Am Soc Nephrol. 2018;29:949–960. 97. Siegerist F, Ribback S, Dombrowski F, et al. Structured illumination microscopy and automatized image processing as a rapid diagnostic tool for podocyte effacement. Sci Rep. 2017;7:11473. 98. Fu Y, Zhu J-Y, Zhang F, et al. Comprehensive functional analysis of Rab GTPases in Drosophila nephrocytes. Cell Tissue Res. 2017;368: 615–627. 99. Zhu J-Y, Fu Y, Richman A, et al. A personalized model of COQ2 nephropathy rescued by the wild-type COQ2 allele or dietary coenzyme Q10 supplementation. J Am Soc Nephrol. 2017;28:2607–2617. 100. Lee HW, Khan SQ, Faridi MH, et al. A podocyte-based automated screening assay identifies protective small molecules. J Am Soc Nephrol. 2015;26:2741–2752. 101. Fan X, Yang H, Kumar S, et al. SLIT2/ROBO2 signaling pathway inhibits nonmuscle myosin IIA activity and destabilizes kidney podocyte adhesion. JCI Insight. 2016;1:e86934. 102. Wei C, Li J, Adair BD, et al. uPAR isoform 2 forms a dimer and induces severe kidney disease in mice. J Clin Invest. 2019;130: pii: 124793. 103. Spinale JM, Mariani LH, Kapoor S, et al. A reassessment of soluble urokinase-type plasminogen activator receptor in glomerular disease. Kidney Int. 2015;87:564–574. 104. Wang X, Dande RR, Yu H, et al. TRPC5 does not cause or aggravate glomerular disease. J Am Soc Nephrol. 2018;29:409–415. 105. Sonneveld R, Hoenderop JG, Isidori AM, et al. Sildenafil prevents podocyte injury via PPAR-g-mediated TRPC6 inhibition. J Am Soc Nephrol. 2017;28:1491–1505.

861